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Nr2e1 Deficiency Augments Palmitate-Induced Oxidative Stress in Beta Cells.

Shi X, Deng H, Dai Z, Xu Y, Xiong X, Ma P, Cheng J - Oxid Med Cell Longev (2015)

Bottom Line: At the molecular level, Nr2e1 deficiency augments palmitate-induced oxidative stress.Nr2e1 deficiency also resulted in decreases in antioxidant enzymes and expression level of Nrf2.Together, this study indicated a potential protective effect of Nr2e1 on beta cells, which may serve as a target for the development of novel therapies for diabetes.

View Article: PubMed Central - PubMed

Affiliation: Department of Endocrinology, Zhongnan Hospital of Wuhan University, Wuhan, Hubei 430071, China.

ABSTRACT
Nuclear receptor subfamily 2 group E member 1 (Nr2e1) has been regarded as an essential regulator of the growth of neural stem cells. However, its function elsewhere is unknown. In the present study, we generated Nr2e1 knockdown MIN6 cells and studied whether Nr2e1 knockdown affected basal beta cell functions such as proliferation, cell death, and insulin secretion. We showed that knockdown of Nr2e1 in MIN6 cells resulted in increased sensitivity to lipotoxicity, decreased proliferation, a partial G0/G1 cell-cycle arrest, and higher rates of apoptosis. Moreover, Nr2e1 deficiency exaggerates palmitate-induced impairment in insulin secretion. At the molecular level, Nr2e1 deficiency augments palmitate-induced oxidative stress. Nr2e1 deficiency also resulted in decreases in antioxidant enzymes and expression level of Nrf2. Together, this study indicated a potential protective effect of Nr2e1 on beta cells, which may serve as a target for the development of novel therapies for diabetes.

No MeSH data available.


Related in: MedlinePlus

Knockdown of Nr2e1 results in reduced expression of Nr2e1 in MIN6 cells. (a) mRNA expression of Nr2e1 in MIN6 cells transduced with shRNA lentivirus targeted against mouse Nr2e1 (MIN6-sh1, MIN6-sh2, and MIN6-sh3) or scrambled nontarget negative control (MIN6-shc). (b) Reduced protein expression of Nr2e1 MIN6-sh1, MIN6-sh2, and MIN6-sh3 cells. Results are means ± SD for three observations. ∗P < 0.05. All comparisons with MIN6-shc cells.
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fig1: Knockdown of Nr2e1 results in reduced expression of Nr2e1 in MIN6 cells. (a) mRNA expression of Nr2e1 in MIN6 cells transduced with shRNA lentivirus targeted against mouse Nr2e1 (MIN6-sh1, MIN6-sh2, and MIN6-sh3) or scrambled nontarget negative control (MIN6-shc). (b) Reduced protein expression of Nr2e1 MIN6-sh1, MIN6-sh2, and MIN6-sh3 cells. Results are means ± SD for three observations. ∗P < 0.05. All comparisons with MIN6-shc cells.

Mentions: We downregulated Nr2e1 protein levels by transduction with lentiviral vectors expressing shRNA that specifically targets Nr2e1 mRNA. Three shRNA sequences targeted to different sites of Nr2e1 mRNA were used, and the transduced MIN6 cells were named MIN6-sh1, MIN6-sh2, and MIN6-sh3. A lentiviral vector carrying a scrambled shRNA sequence was used to create transduced control cells (MIN6-shc). Downregulation of Nr2e1 was confirmed by RT-PCR showing reductions of Nr2e1 mRNA levels by 28% (sh1), 69% (sh2), and 54% (sh3) (Figure 1(a)). A further confirmation was obtained by the western blot analyses showing a decrease of Nr2e1 protein levels by 25%, 71%, and 43%, respectively (Figure 1(b)). Since the sh2 and sh3 target sequences are more effective, they were used for later experiments.


Nr2e1 Deficiency Augments Palmitate-Induced Oxidative Stress in Beta Cells.

Shi X, Deng H, Dai Z, Xu Y, Xiong X, Ma P, Cheng J - Oxid Med Cell Longev (2015)

Knockdown of Nr2e1 results in reduced expression of Nr2e1 in MIN6 cells. (a) mRNA expression of Nr2e1 in MIN6 cells transduced with shRNA lentivirus targeted against mouse Nr2e1 (MIN6-sh1, MIN6-sh2, and MIN6-sh3) or scrambled nontarget negative control (MIN6-shc). (b) Reduced protein expression of Nr2e1 MIN6-sh1, MIN6-sh2, and MIN6-sh3 cells. Results are means ± SD for three observations. ∗P < 0.05. All comparisons with MIN6-shc cells.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4663339&req=5

fig1: Knockdown of Nr2e1 results in reduced expression of Nr2e1 in MIN6 cells. (a) mRNA expression of Nr2e1 in MIN6 cells transduced with shRNA lentivirus targeted against mouse Nr2e1 (MIN6-sh1, MIN6-sh2, and MIN6-sh3) or scrambled nontarget negative control (MIN6-shc). (b) Reduced protein expression of Nr2e1 MIN6-sh1, MIN6-sh2, and MIN6-sh3 cells. Results are means ± SD for three observations. ∗P < 0.05. All comparisons with MIN6-shc cells.
Mentions: We downregulated Nr2e1 protein levels by transduction with lentiviral vectors expressing shRNA that specifically targets Nr2e1 mRNA. Three shRNA sequences targeted to different sites of Nr2e1 mRNA were used, and the transduced MIN6 cells were named MIN6-sh1, MIN6-sh2, and MIN6-sh3. A lentiviral vector carrying a scrambled shRNA sequence was used to create transduced control cells (MIN6-shc). Downregulation of Nr2e1 was confirmed by RT-PCR showing reductions of Nr2e1 mRNA levels by 28% (sh1), 69% (sh2), and 54% (sh3) (Figure 1(a)). A further confirmation was obtained by the western blot analyses showing a decrease of Nr2e1 protein levels by 25%, 71%, and 43%, respectively (Figure 1(b)). Since the sh2 and sh3 target sequences are more effective, they were used for later experiments.

Bottom Line: At the molecular level, Nr2e1 deficiency augments palmitate-induced oxidative stress.Nr2e1 deficiency also resulted in decreases in antioxidant enzymes and expression level of Nrf2.Together, this study indicated a potential protective effect of Nr2e1 on beta cells, which may serve as a target for the development of novel therapies for diabetes.

View Article: PubMed Central - PubMed

Affiliation: Department of Endocrinology, Zhongnan Hospital of Wuhan University, Wuhan, Hubei 430071, China.

ABSTRACT
Nuclear receptor subfamily 2 group E member 1 (Nr2e1) has been regarded as an essential regulator of the growth of neural stem cells. However, its function elsewhere is unknown. In the present study, we generated Nr2e1 knockdown MIN6 cells and studied whether Nr2e1 knockdown affected basal beta cell functions such as proliferation, cell death, and insulin secretion. We showed that knockdown of Nr2e1 in MIN6 cells resulted in increased sensitivity to lipotoxicity, decreased proliferation, a partial G0/G1 cell-cycle arrest, and higher rates of apoptosis. Moreover, Nr2e1 deficiency exaggerates palmitate-induced impairment in insulin secretion. At the molecular level, Nr2e1 deficiency augments palmitate-induced oxidative stress. Nr2e1 deficiency also resulted in decreases in antioxidant enzymes and expression level of Nrf2. Together, this study indicated a potential protective effect of Nr2e1 on beta cells, which may serve as a target for the development of novel therapies for diabetes.

No MeSH data available.


Related in: MedlinePlus